Genotoxic activity of
hexavalent chromium (
chromate) results from its reductive activation inside the cell.
Cr(VI) metabolism in vivo is primarily driven by ascorbate (Asc) but in cultured cells by
glutathione (GSH). Given the common use of cultured cells for mechanistic studies, it is important to establish whether
Cr(VI) activated by Asc and GSH displays the same genotoxic properties. Using 2',7'
dichlorofluorescin (DCFH) as a redox sensitive probe, we found that Asc-dependent reduction of
Cr(VI) in vitro under physiological conditions generated 25-80 times lower yields of
oxidants compared to GSH. When both reducers were present, Asc dominated
Cr(VI) metabolism and inhibited DCFH oxidation. Consistent with the findings in defined chemical reactions, restoration of physiological levels of Asc in human lung H460 cells led to the loss of their
hypersensitivity to clonogenic killing by
Cr(VI) in the presence of
methoxyamine, which inhibits base excision repair of oxidative DNA damage. Despite suppressed oxidative damage, Asc-containing cells formed a large number of
DNA double-strand breaks after exposure to a dose of
Cr(VI) corresponding to the
drinking water standard of 100 ppb. Our results indicate that Asc-driven metabolism of
Cr(VI) shifts its genotoxicity toward nonoxidative mechanisms.